Pharmaceutical composition for treating rheumatism and the preparation thereof

ABSTRACT

The invention has brought to light a kind of antirheumatic and its preparation, which was made from  Tripterygium hypoglaucum  (Levl.) Hutch,  Epimedium brevicornum  Maxim,  Lycium barbarum  L, and  Cuscuta chinensis  Lam. The invented medicine has the merits of prominent effect, mild side reaction and convenient administration.

THE FIELD OF THE INVENTION

The invention is about a medicine which is used to treat rheumatism andits preparation.

THE BACKGROUND OF THE INVENTION

It is believed that the rheumatoid arthritis (RA) is refractory andabout 18,000,000 RA patients have been disabled because of this disease.The medicine reseach for curing RA has continued about a century.Aspirin is the first medicine which is widely used to treat RA. Themedicine to treat RA can be divided into 2 kinds: non-steroidalanti-inflammatory drugs (NSAIDs) and immunosuppressive agent. NSAIDsincludes cyclophthasine, antinfan and adrenal cortex hormone. Theclinical researchs have proved the effectiveness of NSAIDs. Theimmuneosuppressive agent includes methotrexate, cyclophosphane,penicillamine and et al. The immunoregulation has become one of theimportant theropies in the recent years. But all the medicines which areused to treat rheumatism have serious side-effect. The medicine whichcan treat rheumatism effectively and non-poisonously hasn't invented bynow.

There are 3 directions in the research of antirheumatic should beemphasized. The first direction is NSAIDs and cytokine-antagon, such asrecombined soluble TNF α ntagon, IL-1 inhibitor and PAF inhibitor. Thesecond direction is the new immunosuppressive agent and immunomodulator,such as cyclosporin A. The third direction is the compound medicines.

In the TCM, the research on the “arthralgia disease” (equals to thedefinition of rheumatism in the modern medicine) can be traced back tothe Han dynasty more than 1,500 years ago. Three prescriptions: “Ma XingShi Gan decoction”, “Fangji Hangqi decoction” and “Wutong decoction”,which is used to treat “Bi Zheng” were recorded in the medicine classics“Shanghan Lun” wroted by the famous doctor Zhang Zhongjing at that time.Gelsemium elegan s Bentll is a kind of wild plant in Sichuang provinceand it has been proved effective to treat rheumatism in a clinicalresearch carried at the local area. But the further study found that ithad a serious side-effect on the reproduction organs and some otheruncontrollable problem.

The treatment of “arthralgia disease” by the method of TCM has reached ahigh level by numerous doctors' development in so long a history. Bynow, there are many effective prescriptions and herbs. There are morethan 80 kinds of herbs and 29 kinds of patent medicines recorded inChina pharmacopoeia 1995 edition and 2000 edition. But there are stillmany problems for example: {circle over (1)} the effect is not goodenough in treating the serious arthralgia disease such as rheumatoidarthritis; {circle over (2)} the dosage forms are not fit for the modernlife. {circle over (3)} some medicine has good effect, but theside-effect is serious too, such as the extract of triperygiumwilfordii. So that it is necessary to develop thehighly-effective-lowly-noxious and convenient for administrationantirheumatic medicine. This medicine should have the similar effect andthe lower side effect to the artificial antirheumatic medicine.

THE CONTENT OF THE INVENTION

The invention is to supply an antirheumatic, which ishighly-effective-lowly-noxious and convenient for administration, andits preparation thereof.

The invented medicne's technical proposal is realized by using the crudeherbs as following:

-   -   Tripterygium hypoglaucum (Levl.) Hutch.    -   Epimedium brevicornum Maxim.    -   Lycium barbarum L.    -   Cuscuta chinensis Lam. (or Cuscuta australis R. Br.)

The invented medicine is made from the crude herbs above.

The material to produce the invented medicine can be combined on severalways. The tripterygium hypoglaucum (Levl.) Hutch. is the necessary herb,one or two or three of the other three herbs can be added to made thematerial.

One of the optimal crude herbs rate of the material is as following:Tripterygium hypoglaucum (Levl.) Hutch. 1-4 weightinweight Epimediumbrevicornum Maxim. 1-4 weightinweight Lycium barbarum L. 1-4weightinweight Cuscuta chinensis Lam. 1-4 weightinweight

The other optimal crude herbs rate of the material is as following:Tripterygium hypoglaucum (Levl.) Hutch. 2 weightinweight Epimediumbrevicornum Maxim. 2 weightinweight Lycium barbarum L. 1 weightinweightCuscuta chinensis Lam. 1 weightinweight

The third optimal crude herbs rate of the material is as following:Tripterygium hypoglaucum (Levl.) Hutch. 1-4 weightinweight Epimediumbrevicornum Maxim. 1-4 weightinweight

The fourth optimal crude herbs rate of the material is as following:Tripterygium hypoglaucum (Levl.) Hutch. 2 weightinweight Epimediumbrevicornum Maxim. 2 weightinweight

The fifth optimal crude herbs rate of the material is as following:Tripterygium hypoglaucum (Levl.) Hutch. 1-4 weightinweight Epimediumbrevicornum Maxim. 1-4 weightinweight Lycium barbarum L. 1-4weightinweight

The sixth optimal crude herbs rate of the material is as following:Tripterygium hypoglaucum (Levl.) Hutch. 2 weightinweight Epimediumbrevicornum Maxim. 2 weightinweight Lycium barbarum L. 1 weightinweight

The seventh optimal crude herbs rate of the material is as following:Tripterygium hypoglaucum (Levl.) Hutch. 1-4 weightinweight Epimediumbrevicornum Maxim. 1-4 weightinweight Cuscuta chinensis Lam. 1-4weightinweight

The eighth optimal crude herbs rate of the material is as following:Tripterygium hypoglaucum (Levl.) Hutch. 2 weightinweight Epimediumbrevicornum Maxim. 2 weightinweight Cuscuta chinensis Lam. 1weightinweight

The content of the icariine (C₃₃H₄₀O₁₅) in the medicine combinationsabove can not be less than 2.0 mg.

The optimal crude herbs rate of the material can be the other way asfollowing: Tripterygium hypoglaucum (Levl.) Hutch. 1-4 weightinweightLycium barbarum L. 1-4 weightinweight And/or Cuscuta chinensis Lam. 1-4weightinweight

The optimal crude herbs rate of the material can be another way asfollowing: Tripterygium hypoglaucum (Levl.) Hutch. 2 weightinweightLycium barbarum L. 1 weightinweight And/or Cuscuta chinensis Lam. 1weightinweight

The crude herbs are prepared on the rate and then they can be made intoany dosage forms used in the clinic, such as the bolus form, the powderforms, the ointment forms, the tablet forms, the sofe or hard capsuleforms, the granule forms, the injection forms and so on.

The preparation method of the invented medicine is as following:

The crude herbs are prepared on the weight rate: Tripterygiumhypoglaucum (Levl.) Hutch 1-4 weightinweight Epimedium brevicornum Maxim1-4 weightinweight Lycium barbarum L 1-4 weightinweight Cuscutachinensis Lam 1-4 weightinweight

The Tripterygium hypoglaucum (Levl.) Hutch. and Epimedium brevicornumMaxim are smashed. Then the powders are decocted by water for 2˜4 timesseparately. The Lycium barbarum L and Cuscuta chinensis Lam are soakedin the hot water (80˜95° C.) for 1˜3 times separately. The decoctedfluid and the immersion fluid of the herbs are collected and added tothe correspondent macroscopicvoid adsorbent resins column separately.After the adsorption, the columns are washed until the flushing liquortunns clear. Then the columns are eluted by 60%-80% alcohol. The elutingliquors are collected from its color turning deep till the color turningvery weak. Then the alcohol in the upper part of the column is pushedout by high pressure water and mixed with the eluting liquor. The mixedeluting liquor is 3˜8 times heavy of the correspondent crude herb. Allthe 4 eluting liquors are recycled and condensed to the specific density1.10 separately. The condensed liquors are dehydrated by spry drying toget the extract of the crude herbs. The 4 kind of extracts are mixeduniformly to be made into any dosage forms that needed by the clinic.

The optimal preparation method of the invented medicine is as following:

The crude herbs are prepared on the weight rate: Tripterygiumhypoglaucum (Levl.) Hutch 2 weightinweight Epimedium brevicornum Maxim 2weightinweight Lycium barbarum L 1 weightinweight Cuscuta chinensis Lam1 weightinweight

The Tripterygium hypoglaucum (Levl.) Hutch. Is smashed. Then the powderis added with 13, 10, 10 folds weight of the water to decoct for 3times. Each time is 1 hour. The Epimedium brevicornum Maxim is cut topiece. Then the herb pieces is added with 15, 10, 10 folds weight of thewater to decoct for 3 times. Each time is 1 hour. The Lycium barbarum Lis smashed to crude powder and soaked in the hot water (80° C., 20 foldweight of the crude herb) for 3 times. Each time is 1 hour. The Cuscutachinensis Lam is smashed to crude powder and soaked in the hot water(80° C., 31 fold weight of the crude herb) for 3 times. Each time is 1hour. The decocted fluid and the immersion fluid of the herbs arefiltrated separately and added to the correspondent macroscopicvoidadsorbent resins column JD-1 (WLD). After the adsorption, the columnsare eluted by 70% alcohol. The eluting liquors are collected from itscolor turning deep till the color turning very weak. The alcohol isrecycled from the eluting liquor. Then the rest liquor is condensed anddehydrated to get the extract powder. The 4 kind of extract powders aremixed uniformly to be made into any dosage forms that needed by theclinic.

The invented medicine can be prepared on the method as following:

The crude herbs are prepared on the weight rate recorded before. TheTripterygium hypoglaucum (Levl.) Hutch. and Epimedium brevicornum Maximare cut into pieces. The Lycium barbarum L and Cuscuta chinensis Lam arecrushed or not. The 4 kind of herbs are extracted in the 0˜95% alcoholat 10˜98° C. for 1˜4 times separately or together. The extracted liquorsare mixed or not. Then the extracted liquors are condensed, dehydrated,smashed and mixed uniformly. The mixed powder can be made into anydosage form needed in the clinic.

The invented medicine can be made from the effective constituents of the4 herbs.

The effective constituents of Epimedium brevicornum Maxim are icariine,icariside I, icariside II, and Icariin A. The effective constituents ofTripterygium hypoglaucum (Levl.) Hutch. Are diterpenes, triterpenes andalkaloids compound. The effective constituents of Lycium barbarum L andCuscuta chinensis Lam are both flavone.

So that the crude herb Epimedium brevicornum Maxim can be replaced byone or more kinds of the effective constituents of itself, such asicariine, icariside I, icariside II, and Icariin A. The crude herbTripterygium hypoglaucum (Levl.) Hutch. Can be replaced by one or morekinds of the effective constituents of itself, such as diterpenes,triterpenes and alkaloids compound. While the Lycium barbarum L andCuscuta chinensis Lam can be replaced by flavone.

It has been proved by the pharmacodynamics research that the inventedmedicine (Fengshiping Capsule) could inhibit the primary and secondaryinjury adjuvant arthritis (AA). It could inhibit the delayedhypersensitivity (DTH) in the ear of the mouse caused by the2,4dinitrofluorobenzene (DNFB). It could inhibit the antibody produce ofthe hemolysin and the activity of the IL-1, IL-2, IL-6 and TNF in themacrophage and splenocyte. The Fengshiping Capsule could inhibit thelymphocyte transformation induced by the ConA. It could inhibit the CD₄,CD₈ cells remarkably, especially CD₄ cells, but the rate of CD₄/CD₈ wasnot affected very much. There was a remarkable linear relationshipbetween the dosage and the effect. 12˜18 g (crude medicine)/kg was theminimum effective dose. The invented medicine could inhibit the activityof the NK cells. In the effective dose, Fengshiping Capsule did notcause the atrophy of the important immune organs such as thymus andspleen, and did not inhibit the phagocytic activity of the macrophage.

The invented medicine had a remarkable antiinfalmmatory action. It couldinhibit the over penetrating condition of the capillary in the mouse'sabdominal cavity caused by the ethanoic acid injected. It could improvethe swelling in the ear of the mouse caused by the croton oil. It couldinhibit the pleuritis in the mouse and the assembling of the WBC to theCMC cyst in the rat induced by the carrageenan. But the inventedmedicine couldn't inhibit the rat's foot swelling induced by thecarrageenan and the granuloma caused by the tampon obviously. TheFengshipng Capsule could inhibit the body-twist reaction caused by theethanoic acid in the mouse remarkably.

EXPERIMENTAL EXAMPLE 1 The Effect on the Adjuvant Arthritis (AA)

1.1 The Preventing Effect on the AA of the Invented Medicine

72 isogenous SD rats of the same batch, half male and half female,180˜220 g weight, were divided randomly into 6 groups. Each group has 12rats. Every 6 rats lived in a cage. The perimeter of the doubleanklejoints and the feet of the rat were measured accurately andrecorded as the normal value. All the rats were drenched by the samevolume of the invented medicine on the correspondent concentration orthe solution of the Xihuangqi by the gastic injection. 1 hour later, allthe rats were injected with 0.1 ml Freund's complete adjuvant (FCA)under the skin of the left postpedes. In the next 30 days, all the ratswere drenched with the correspondent medicine once a day on the samedosage. And in these days, the rats were measured of the perimeters ofthe double anklejoints and the feet once a day. In this experiment, theswelling degree (Δcm) equaled to the difference value of the perimetersmeasured after the FCA injection and before the FCA injection. (See theresult in table 1.1 and 1.2) At the end of the experiment, the majororgans of the rats were weighted. (See the table 1.3, 1.4) TABLE 1.1 Theeffect of the Fengshiping on the swelling degree of the left anklejointand foot after the injection of FCA in the rat AA model ({overscore (X)}± S) Dose Swelling degree (Δcm) Group (g/kg) 1 d 2 d 3 d 9 d 12 d 14 d16 d Control — 0.69 ± 0.17 0.69 ± 0.12 0.92 ± 0.18 0.84 ± 0.41 1.10 ±0.30 1.65 ± 0.68 2.10 ± 0.55 Fengshiping 7.5 0.74 ± 0.12 0.66 ± 0.0740.83 ± 0.13 0.77 ± 0.27 1.11 ± 0.45 1.34 ± 0.53 1.91 ± 0.61 Fengshiping15 0.80 ± 0.24 0.62 ± 0.13 0.76 ± 0.18 0.49 ± 0.17* 0.73 ± 0.34* 1.00 ±0.48* 1.38 ± 0.67* Fengshiping 30 0.75 ± 0.19 0.67 ± 0.19 0.87 ± 0.280.63 ± 0.22 0.73 ± 0.34* 0.82 ± 0.43** 1.05 ± 0.53** Tripterygium 5 0.72± 0.11 0.68 ± 0.16 0.91 ± 0.18 0.66 ± 0.23 0.88 ± 0.29 1.03 ± 0.36* 1.37± 0.33* hypoglaucum (Levl.) Hutch. prednisone 0.01 0.64 ± 0.14 0.64 ±0.16 0.50 ± 0.26 0.46 ± 0.25 0.72 ± 0.46* 0.87 ± 0.46** 1.28 ± 0.69*Dose Swelling degree(Δcm) Group (g/kg) 18 d 20 d 22 d 24 d 26 d 28 dControl — 2.18 ± 0.44 2.05 ± 0.46 2.00 ± 0.46 2.04 ± 0.57 1.92 ± 0.651.83 ± 0.67 Fengshiping 7.5 1.74 ± 0.73 1.81 ± 0.55 1.81 ± 0.52 1.77 ±0.55 1.65 ± 0.55 1.55 ± 0.49 Fengshiping 15 1.32 ± 0.59** 1.28 ± 0.58**1.34 ± 0.61* 1.33 ± 0.67* 1.20 ± 0.64* 1.08 ± 0.58** Fengshiping 30 0.95± 0.50** 0.87 ± 0.51** 0.95 ± 0.54** 0.89 ± 0.59** 0.90 ± 0.57** 0.86 ±0.51** Tripterygium 5 1.47 ± 0.43** 1.50 ± 0.43** 1.49 ± 0.43* 1.42 ±0.53* 1.40 ± 0.56* 1.32 ± 0.57 hypoglaucum (Levl.) Hutch. prednisone0.01 1.18 ± 0.7**6 1.03 ± 0.67** 1.05 ± 0.69* 0.90 ± 0.64** 0.86 ±0.65** 0.85 ± 0.59**Comparing to the control group *P < 0.05, **P < 0.01 (the signs have thesame meaning in the following tables)

TABLE 1.2 The effect of the Fengshiping on the swelling degree of theleft anklejoint and footafter the injection of FCA in the rat AA model({overscore (X)} ± S) Dose Swelling degree (Δcm) Group (g/kg) 2 d 9 d 12d 14 d 16 d 18 d Control — 0.14 ± 0.05 0.06 ± 0.10 0.34 ± 0.36 0.80 ±0.52 1.43 ± 0.67 1.36 ± 0.61 Fengshiping 7.5 0.18 ± 0.06 0.10 ± 0.140.26 ± 0.36 0.82 ± 0.52 1.31 ± 0.64 1.28 ± 0.71 Fengshiping 15 0.15 ±0.08 0.02 ± 0.06 0.13 ± 0.10* 0.37 ± 0.31* 0.90 ± 0.56* 0.79 ± 0.60*Fengshiping 30 0.18 ± 0.09 0.06 ± 0.06 0.16 ± 0.08* 0.29 ± 0.20** 0.49 ±0.41** 0.33 ± 0.29** Tripterygium 5 0.16 ± 0.07 0.01 ± 0.07 0.11 ± 0.100.44 ± 0.19** 0.87 ± 0.56* 0.84 ± 0.67* hypoglaucum (Levl.) Hutch.prednisone 0.01 0.20 ± 0.06 0.08 ± 0.08 0.21 ± 0.16 0.44 ± 0.43 0.99 ±0.63 0.84 ± 0.74* Dose Swelling degree (Δcm) Group (g/kg) 20 d 22 d 24 d26 d 28 d Control — 1.28 ± 0.57 1.38 ± 0.64 1.35 ± 0.75 1.20 ± 0.78 1.12± 0.63 Fengshiping 7.5 1.33 ± 0.71 1.31 ± 0.73 1.27 ± 0.73 1.16 ± 0.731.07 ± 0.65 Fengshiping 15 1.74 ± 0.57* 1.92 ± 0.61* 0.95 ± 0.64* 0.88 ±0.58* 1.83 ± 0.55 Fengshiping 30 0.27 ± 0.30** 0.34 ± 0.31** 0.32 ±0.33** 0.31 ± 0.32** 0.34 ± 0.32** Tripterygium 5 0.82 ± 0.65* 0.89 ±0.70* 0.80 ± 0.67* 0.83 ± 0.68 0.75 ± 0.69 hypoglaucum (Levl.) Hutch.prednisone 0.01 0.82 ± 0.72* 0.79 ± 0.74* 0.75 ± 0.67** 0.68 ± 0.64*0.71 ± 0.67

1.3 The effect of the Fengshipng on the body weight of the AA rats (X ±S) Body weight change(g) Dose BW at 1 month Group (g/kg) Initiative BWlater BW change Control — 228 ± 34 231 ± 52 3 Fengshiping 7.5 229 ± 34220 ± 46 −9 Fengshiping 15 223 ± 40 232 ± 34 9 Fengshiping 30 224 ± 37256 ± 60 32 Tripterygium 5 226 ± 45 230 ± 43 4 hypoglaucum (Levl.)Hutch. prednisone 0.01 264 ± 55 244 ± 31 −21

TABLE 1.4 The effect of the Fengshiping on the organ weight of theimmune system in the AA rats (prevention experiment)(X ± S) Dose Organindex [(organ weight/body weight)/100] Group (g/kg) Liver Spleen Thymusadrenal gland Control — 3.92 ± 0.65 0.34 ± 0.10 0.098 ± 0.040 0.027 ±0.01 Fengshiping 7.5 3.73 ± 0.29 0.31 ± 0.09 0.078 ± 0.038 0.027 ± 0.008Fengshiping 15 3.48 ± 0.32 0.38 ± 0.10 0.100 ± 0.034 0.023 ± 0.005Fengshiping 30 3.38 ± 0.28* 0.44 ± 0.12* 0.100 ± 0.032 0.022 ± 0.007Tripterygium 5 3.21 ± 0.30** 0.36 ± 0.05 0.052 ± 0.011** 0.026 ± 0.009hypoglaucum (Levl.) Hutch. prednisone 0.01 3.04 ± 0.20** 0.32 ± 0.080.050 ± 0.060** 0.020 ± 0.004*1.2 The Therapeutic Effect on the AA of the Invented Medicine

50 male SD rats were divided into 5 groups at random. The model buildingwas same to the prevention experiment, but the correspondent medicineswere drenched 13 days after the injection of the FCA. The medicines weredrenched once a day for 2 weeks. The swelling degree (Δcm) was thedifference of the perimeters between the value of first administrationday and the other days. (Se the result in table 1.5, 1.6) The majororgans' weight is showed in table 1.7. TABLE 1.5 The therapeutic effectof Fengshiping on the swelling degree of The left anklejoint and foot inthe AA rats ({overscore (X)} ± S) Dose Swelling degree (Δcm) Group(g/kg) 1 d 2 d 4 d 6 d Control — 1.81 ± 0.27 1.92 ± 0.19 2.12 ± 0.222.16 ± 0.27 Fengshiping 7.5 1.68 ± 0.50 1.64 ± 0.54 1.70 ± 0.57 1.82 ±0.61 Fengshiping 15 1.44 ± 0.41* 1.51 ± 0.36** 1.65 ± 0.34** 1.74 ±0.31** Fengshiping 30 1.50 ± 0.56 1.48 ± 0.41** 1.51 ± 0.44** 1.59 ±0.51** prednisone 0.01 1.78 ± 0.51 1.70 ± 0.51 1.63 ± 0.50* 1.58 ±0.50** Dose Swelling degree (Δcm) Group (g/kg) 8 d 10 d 12 d 14 dControl — 1.92 ± 0.32 1.87 ± 0.34 1.92 ± 0.39 1.78 ± 0.44 Fengshiping7.5 1.67 ± 0.68 1.60 ± 0.71 1.61 ± 0.77 1.58 ± 0.71 Fengshiping 15 1.46± 0.37** 1.48 ± 0.30* 1.28 ± 0.37** 1.22 ± 0.38** Fengshiping 30 1.29 ±0.58** 1.29 ± 0.65** 1.26 ± 0.67** 1.20 ± 0.68* prednisone 0.01 1.27 ±0.46** 1.09 ± 0.54** 0.94 ± 0.50** 0.94 ± 0.42**

TABLE 1.6 The therapeutic effect of Fengshiping on the swelling degreeof the right anklejoint and foot in the AA rats (X ± S) Dose Swellingdegree (Δcm) Group (g/kg) 2 d 4 d 6 d 8 d 10 d 12 d 14 d Control — 0.36± 0.26 0.45 ± 0.25 0.55 ± 0.34   0.47 ± 0.29   0.48 ± 0.25   0.46 ± 0.31  0.40 ± 0.36 Fengshiping 7.5 0.12 ± 0.25 0.34 ± 0.32 0.48 ± 0.41   0.28± 0.38   0.35 ± 0.30   0.30 ± 0.29   0.30 ± 0.35 Fengshiping 15 0.21 ±0.18 0.38 ± 0.27 0.44 ± 0.33   0.21 ± 0.33*   0.19 ± 0.45*   0.06 ±0.31** −0.06 ± 0.34** Fengshiping 30 0.10 ± 0.48 0.06 ± 0.28** 0.11 ±0.24**   0.06 ± 0.27**   0.02 ± 0.39**   0.05 ± 0.38* −0.02 ± 0.41**prednisone 0.01 0.10 ± 0.13* 0.15 ± 0.28* 0.11 ± 0.25** −0.08 ± 0.34**−0.13 ± 0.28** −0.26 ± 0.36** −0.33 ± 0.39**n = 10,comparing with the control group, *P < 0.05, **P < 0.01

TABLE 1.7 The effect of the Fengshiping on the organ weight of theimmune system in the AA rats (X ± S) Dose Organ index [(organweight/body weight)/100] Group (g/kg) Liver Spleen Thymus adrenal glandControl — 0.35 ± 0.23 0.35 ± 0.061 0.073 ± 0.014 0.026 ± 0.0071Fengshiping 7.5 3.21 ± 0.52 0.33 ± 0.091 0.071 ± 0.026 0.024 ± 0.0085Fengshiping 15 3.40 ± 0.54 0.36 ± 0.014 0.067 ± 0.022 0.023 ± 0.0048Fengshiping 30 2.79 ± 0.43 0.32 ± 0.083 0.069 ± 0.029 0.023 ± 0.0072Tripterygium 5 3.92 ± 0.59 0.35 ± 0.100 0.075 ± 0.034 0.027 ± 0.0060hypoglaucum (Levl.) Hutch. prednisone 0.01 3.52 ± 0.35 0.28 ± 0.047* 0.05 ± 0.011**  0.02 ± 0.0043*

The data showed in the table 1.1, 1.2, 1.3, 1.5 and 1.6 proved that theFengshiping could strongly inhibit the primary and secondary injurycaused by FCA, whenever the medicine was drenched at the beginning ofthe FCA injection or 2 weeks after the FCA injection. The experimentsproved that the Fengshiping had both the preventing and the therapeuticeffect. By comparing the effect of Fengshiping on the swelling degree inthe anklejoint and foot, we found that the Fengshiping could inhibit thespecific immunoswelling in the anklejoint better than the nonspecificimmunoswelling in the foot of rats. This result indicated that the maineffect of Fengshiping was inhibiting the immunity inflammatory reaction.

The data in the table 1.3, 1.4 and 1.7 showed that the AA rats had noobvious BW increase during the period of the experiment. In the groupdrenched of the Fengshiping on the effective dosage, the rats still hadBW increase. In the groups of prednisone and preventing, the BW of ratshad decreased, while the thymus and adrenal gland were atrophied. In thegroup of tripterygium hypoglaucum (Levl.) Hutch, the thymus had thrinkedyet. But in the 3 groups drenched with the different dosage ofFengshiping, on atrophy of the thymus and adrenal gland were observed.

1.3 The Pathologic Change of the AA after the Treatment of the InventedMedicine in Rats

45 SD rats, 180±20 g weight, were divided into 6 groups. After the AAcaused by FCA appeared, all the rats were drenched with Fengshipingsolution by gastic injection for 5 days once a day. 1 hour after thelast administration, the joint index of the rats was measured andcalculated. The secondary injuried postpedes' joints on the opposite ofthe FCA injectiont were taken off and soaked in the formaldehyde. Afterthe tissues were HE tinted, the pathological change of the synovium andcartilage were observed and recorded. The data were showed in table 1.8.1.8 The effect of Fengshiping on the AA joint index in the rats (X ± S)Dose Rat Group (g/kg) number Joint index Control — 8 0** AA model — 76.2 ± 0.49  Fengshiping 7.5 9 4.86 ± 0.90** Fengshiping 15 7 4.71 ±0.95** Fengshiping 30 7 4.56 ± 1.13** Glucosidorum Tripterygll Totorum0.006 7 4.57 ± 0.79**Comparing with the model group **P < 0.01

The joint index was the sum of the inflammatory scores of the fourlimbs. According to the degree of inflammatory, each limb was evaluatedon the criteria as following: normal (0), red without swelling (1), redand swelling (2), seriously swelling (3), deforming and tetanus (4).

Observed from the microscope, the joint synovial membranes of the ratposterior limb were hyperplasia in the model group; and the collagenfiber had increased; there was infiltration of lymphocytes and plasmacells in the tissue. The obvious granuloma had formed. The synovialcells had degenerated and the cytochylema had been tinted red; thecaryon had been pycnosis; the epithelium had exfoliated in some part ofthe synovial membrane. The cartilage turned atrophy; the surface of itwas rough and some of the chondrocytes had proliferated lightly.

After the treatment of the Fengshiping, the inflammation of the jointsynovial membrane was inhibited, more collogen fiber was produced; lesssynovial cells exfoliated; the cells on the surface of the cartilage hadproliferated and the surface had turned smooth. The cartilage was on therecovering condition.

EXPERIMENTAL EXAMPLE 2

The effect of Fengshiping on the delayed hypersensitivity reaction (DTH)caused by 2,4-DNFB in the ear of the mouse 50 NIH mice, half male andhalf female, were divided into 5 groups. Each mouse was led tohypersensitivity reaction by using the 1% DNFB acetone solution on thedosage of 0.025 ml at the right place of the abdomen where the pilus hadbeen cut yet. Using the same solution on the same place was to enhancethe hypersensitivity reaction on the third day. On the fifth day, allthe mice were smeared with the 1% DNFB edible oil solution at the mice'sright ears on the dosage of 0.01 ml each. 24 hours later, all the micewere killed. The mouse's 2 ears were weighted by the torsion balance andthe difference of the 2 ears was recorded as the DTH degree caused bythe DNFB. The experiment was carried out on the different immune andadministration processes.

2.1 The Effect on the DTH by the Full Course Administration

The immune and administration processes is as following:

TABLE 2.1 The effect of Fengshiping on the DTH caused by DNFB in the NIHmouse (X ± S) dose Administration time Percent of inhibition group(g/kg) (day) Mice number Percent of ear swelling (%) P value control 1034.20 ± 3.77 Fengshiping 27 0˜5 10 26.24 ± 3.34 23.3 <0.01 Fengshiping40 0˜5 10 12.99 ± 4.96 62.0 <0.01 Fengshiping 60 0˜5 10 10.43 ± 7.5369.5 <0.01 cortisumman 0.003 0˜5 10 13.93 ± 4.41 59.3 <0.01 control 1042.43 ± 5.28 Fengshiping 40 −2˜0   10  31.50 ± 10.52 25.0 <0.01Fengshiping 40 −2˜2   10 30.88 ± 7.92 27.2 <0.01 Fengshiping 40 −2˜5  10  21.07 ± 4.62* 50.3 <0.01 Fengshiping 40 5˜6 10 32.00 ± 9.37 41.7<0.01 cyclophosphane 0.05 −2˜2   10  39.40 ± 10.78 8.1 >0.05cyclophosphane 0.05 −2˜0   10 37.47 ± 6.71 11.7 >0.05 control 10 38.50 ±4.67 cyclophosphane 0.1 × 3 0, 2, 4 day once a day 10 23.00 ± 7.65 40.3<0.01 cyclophosphane 0.25 −3 d 10 41.84 ± 7.75 −8.7 Fengshiping 60 0˜410  27.20 ± 10.20 29.4 <0.01 cyclophosphane + Fengshiping 0.25 + 60 −3,0˜4 10 38.07 ± 6.65 1.1*comparing with the other groups P < 0.05

P < 0.01

According to the data showed in table 2.1, it indicated that theFengshiping had a obvious inhibiting effect on the DTH caused by DNFB.There was a significant relationship between the dosage and the effect.The inhibiting activity increases when the dosage adds. The inhibitingpercent could reach 69.5% on the dosage of 60.9 g/kg.

2.2 The Effect on the DTH of the Different Administration Time

The immune and administration processes and the correspondent resultshad been showed in middle and bottom parts of the table 2.1. Accordingto the results showed in the table 2.1, all the administration wayscould significantly inhibit the DTH of the mouse in spite of theadministration beginning from the 2 days before the sensitization andending at the sensitization, or beginning from the 2 days before thesensitization and ending 2 days after the sensitization, or beginningfrom the 2 days before the sensitization and ending 5 days after thesensitization, or beginning before the attack and ending after theattack. But the administration way that began 2 days before thesensitization and ended 5 days after the sensitization had the mostpowerful inhibiting activity. It indicated that the Fengshiping couldinhibit the DTH by a compound mechanism that it could inhibit the cellsparticipant in the early period of the DTH, the effector cells in theadvanced period and the cells related to the DTH in the middle period.This mechanism was different from that of the cyclophosphane. On a smalldosage, the cyclophosphane didn't affect the DTH, if its administrationway was beginning from the 2 days before the sensitization and ending atthe sensitization day or 2 days after the sensitization day.

Based on the bottom part of the table 2.1, if a high dosage ofcyclophosphane was drenched to the mouse in one time 3 days before thesensitization, the function of the Th cells would turn sthenic becauseof the powerful inhibition on the Ts cells. The DTH in the mouse wouldbe enhanced. If the cyclophosphane was used with the Fengshiping on thisadministration way, it could lower the inhibiting activity ofFengshiping. This result indicated that the Fengshiping have a differentmachnizm to the cyclophosphane in the control of DTH. The Fengshipingmaybe had a higher activity in inhibiting the TH cells.

EXPERIMENTAL EXAMPLE 3 The Effect on the Humoral Immunity

3.1 The Effect on the Produce of the Hemolysin Antibody Caused by theChick RBC

190 mice, 18-22 g weight, half male and half female, were divided into19 groups. Each mouse was immunized with 5% CRBC solution 0.2 ml. TheFengshiping solutions were drenched at the different times. 7 days afterthe immunization, all the mice were sampled the blood from the eyes.Then the blood samples were diluted and measured the level of thehemolysin antibody. The results were showed in table 3.1, 3.2 and 3.3.TABLE 3.1 The effect of Fengshiping on the produce of the hemolysinantibody in the NIH mouse (X ± S) dose Administration Mouse HemolysinInhibiting group (g/kg) time number value percent (%) P value control 10169.0 ± 62.0  Fengshiping 18 0˜7 10 46.0 ± 15.6 72.8 <0.01 Fengshiping27 0˜7 10 35.4 ± 12.0 79.1 <0.01 Fengshiping 40 0˜7 10 28.2 ± 5.9  83.3<0.01 Fengshiping 60 0˜7 10 16.7 ± 3.0  90.1 <0.01 Tripterygium 13.3 0˜710  121.0 ± 88.0** 28.4 <0.015 hypoglaucum (Levl.) Hutch. cyclophosphane0.02 0˜7 10 35.0 ± 12.0 79.3 <0.01**comparing with the Fengshiping (40 g/kg) group P < 0.01

TABLE 3.2 The effect of Fengshiping on the produce of the hemolysinantibody in the ICR mouse (X ± S) dose Administration Mouse inhibiting Pgroup (g/kg) time number Hemolysin value percent (%) value control — —10 124.70 ± 42.60  Fengshiping 12 0˜7 10 75.00 ± 53.10 39.9 <0.05Fengshiping 18 0˜7 10 45.60 ± 22.70 63.4 <0.01 Fengshiping 27 0˜7 1029.10 ± 22.10 76.8 <0.01 Fengshiping 40 0˜7 10 28.20 ± 5.30  77.4 <0.01Tripterygium 6.0 0˜7 10  143.50 ± 67.90** >0.05 hypoglaucum (Levl.)Hutch. cyclophosphane 0.02 0˜7 10 27.80 ± 6.60  77.9 <0.01**comparing with the Fengshiping (18 g/kg) group P < 0.01

TABLE 3.3 The effect of Fengshiping on the produce of the hemolysinantibody in the ICR mouse (X ± S) dose Administration Mouse HemolysinInhibiting percent P group (g/kg) time number value (%) value control —— 10 256.0 ± 26.0 Fengshiping 18 −7˜7   10 198.0 ± 50.0 22.7 <0.01Fengshiping 18 −3˜7   10 156.0 ± 85.0 39.1 <0.01 Fengshiping 18 0˜7 10 98.0 ± 35.0 61.7 <0.01 cyclophosphane 0.02 0˜7 10 25.0 ± 4.0 90.2 <0.01

According to the data in table 3, it indicated that the Fengshiping hada remarkable inhibiting effect on the produce of the hemolysin antibodyin the different mouse species and this effect would increase along withthe increase of the dosage. There was a certain relationship between thedosage and the effect. The lowest effective dosage was 12 g/kg.Comparing with the same quantity of Tripterygium hypoglaucum (Levl.)Hutch, the Fengshiping had a higher inhibiting activity. Based on thedata in table 3.1, the inhibiting activity of Fengshiping was 2.25 timeshigher than the Tripterygium hypoglaucum (Levl.) Hutch. The inhibitingactivity of Tripterygium hypoglaucum (Levl.) Hutch. On the dosage of13.5 g/kg was weaker than that of the Fengshiping which containing 6g/kg Tripterygium hypoglaucum (Levl.) Hutch).

3.2 The Effect of the Fengshiping on the Humoral Immunity in the AAMouse

The NIH mice, 20±2 g weight, were injected with 0.05 ml FCA under thevola skin of the right postpedes. 3 weeks late the AA model mousebuilded. The model mice were divided into 6 groups randomly and drenchedwith the correspondent medicines for 5 days. At the beginning of theadministration, all the mice were sensitized with 0.5 ml 10% sheep RBC(SRBC). Five days later, all the mice were killed. Their spleens weretaken out and washed by the Hank's liquor to prepare the lymphocytesuspended liquor. The concentration of the cells was adjusted to2×10⁷/ml. 1 ml lymphocyte suspension, ml 0.2% SRBC and 1 ml 1:30addiment were added to one test tube. The tube was put in the water bathat 37° C. for 1 hour. Then the tube was centrifugated at 2000 rpm for 5minutes. The supernatant fluid was separated and tested its spticaldensity at the 415 nm wavelength on the 722 apeotrophotometer. The valuewas the represent of PFC quantity.

The other share of the blood samples got from the sensitized mice wasseparated the serum to test the potency of the antibody. The measureddata were recorded on the way of Log 2 value. (See the data in table3.4) TABLE 3.4 The effect of Fengshiping on the humoral immunity in themouse (X ± S) dose Mouse PFC group (g/kg) number (OD) IgM(Log2) control— 8 0.819 ± 0.013# 6.875 ± 0.641 AA model — 10 0.940 ± 0.019** 7.700 ±0.599* group fengshiping 5 8 0.834 ± 0.012**# 6.875 ± 0.641# fengshiping10 8 0.834 ± 0.012**# 6.750 ± 0.886# fengshiping 20 8 0.830 ± 0.014**#6.375 ± 0.518## Glucosidorum 0.012 10 0.835 ± 0.015**# 6.950 ± 0.597#Tripterygll TotorumComparing with the control group *P < 0.05, **P < 0.01; comparing withthe model group #P < 0.05, ##P < 0.01

According to the table 3.4, the levels of PFC and IgM in the AA mousewere higher than that of the normal mouse. The Fengshiping could lowerthe produce of the PFC and IgM in the AA mouse significantly.

EXPERIMENTAL EXAMPLE 4 The Effect of the FENGSHIPING on the PassiveCutis Anaphylactic Reaction (PCA) in the Rat

The rats were injected with the egg albumn at 10 mg/kg in the muscle. Atthe same time, all the rats were injected with 2×10¹⁰ (0.2 ml)bordetella pertussis in the abdominal cavity for sensitization. 2 weekslater, all the rats were killed to sample the blood. All the bloodsamples were separated for preparing the serum.

60 rats, 150˜200 g, half male and half female, were divided into 6groups at random. In the light narcosis condition induced by aether,each rat was cut the fleece in the back and injected with the 2concentrations of anti-egg-album serum 0.1 ml under the skin at thefairless place. The serums were diluted on the rates of 1:5(d1) and1:10(d2) before the experiment. 48 hours later, all the rats wereattacked by intravenous injecting the 0.5% evans blue normal salinesolution 1 ml which containing 1 mg egg albumin. 20 minutes later, therats were killed by decapitation. The rats' back skin were dissected andturned over. According to the dark and light and area of the blue stainsexudated from the vessels, all the rats were evaluated by severalpeople. The skin stained by the evens blue were scissored and soaked in5 ml 0.1% sodium sulfate acetone (7:3) solution for 48 hours. Then itwas centrifugated to separate the supernatant liquor. The supernateswere measured the optical density at the wavelength 590 nm to calculatethe degree of the PCA reaction and the inhibiting percent. The resultswere shown in table 4. TABLE 4 The effect of Fengshiping on the PCA inrat (X ± S) dose value absorbancy Group (g/kg) d₁ d₂ d₁ d₂ Control —5.60 ± 1.78 2.40 ± 2.46 0.191 ± 0.129 0.096 ± 0.106 Fengshiping 12 7.50± 2.51 4.20 ± 2.49 0.402 ± 0.213* 0.192 ± 0.175 Fengshiping 24 7.10 ±2.13 4.10 ± 1.79 0.310 ± 0.177 0.137 ± 0.099 Fengshiping 48 6.00 ± 1.831.70 ± 1.95 0.121 ± 0.109 0.024 ± 0.026* Tripterygium hypoglaucum 8 6.11± 1.27 2.56 ± 1.67 0.223 ± 0.122 0.074 ± 0.045 (Levl.) Hutch. Ketotifen0.1 2.78 ± 1.64** 0.67 ± 1.41 0.033 ± 0.024** 0.027 ± 0.019*Comparing with the control group *P < 0.05, **P < 0.01

According to the table 4, it indicated that the Fengshiping had a weakeffect on the PCA in the rat. Only on a high dosage, the inhibitingeffect of Fengshiping was obviously different from that of the controlgroup.

EXPERIMENTAL EXAMPLE 5 The Effect of Fengshiping on the Cytokines

5.1 The effect of Fengshiping on the levels of TNF α and IL-2 in themouse.

60 ICR mice, 1822 g, half male and half female, were divided into 6groups at random. Each group was drenched of the correspondent medicinesincluding the different dosages of Fengshiping and the other medicines.The medicines were administrated once a day for 10 days. 24 hours afterthe last administration, the mice were sampled the macrophage and spleencells from the abdominal cavity in the aseptic condition. The sampleswere washed with Hank's liquor for 2 times and non-serum RPIM 1640liquor for 1 time. Then the washed samples were diluted to thesuspension with the 5% FCS-RPMI 1640 at the concentration of 2×10⁸/ml.Then the suspensions were added with 10 ng/ml LPS or the 10 ng/ml ConAand cultured in the 5% CO2 condition for 48 hours at 37° C. Then thecultured suspension were measured the TNF α and IL-2 levels on the usualmethods.

The Measurement of TNF α

The batten was coated by mouse TNF-α monoclonal antibody. The batten wasadded with the cultured supernate on the dose of 50 μl/hole. Then thebatten was put still for 60 minutes at the room temperature. Then thebatten was added with biotin antibody mark at 25° C. for 2 hours. Thenthe enzyme labeled avidin was added into the batten for 30 minutes.After adding the substrate constant for 30 minutes, the batten was addedwith the stop liquor. The mixed liquor was measured the OD value at thewavelength of the 450 nm. The content of the TNF-α (ng/ml) wascalculated on the data of OD value by the method of standard curve.

The Measurement of the IL-2:

The CTLL cells which was on the logarithmic growth phase and whosegrowth depends on the IL-2, were adjusted to the suspension at theconcentration of 1×10⁵/ml with the 5% FCS-RPMI 1640. Then the 96 holecell culturing batten were added with the CTLL cell suspension on thequantity of 100 μl/hole. The supernates were added on the quantity of100 μl/hole and each sample was added to 3 holes. The samples culturedwere compared with the different dilutions of standard rHIL-2 and thecontrol sample (culture fluid) to measure the IL-2. All the samples werecultured in the 5% CO2 for 24 hours at 37° C. 6 hours before the end ofthe culture, all the samples were centrifuged and separated thesupernate. Each hole were taken out 110 μl supernate and added with 10μl MTT. The samples were cultured for 3 hours at 37° C., and thenmeasured the OD at the wavelength 570 nm and 630 nm. The final OD valueof the sample was the difference of OD (570 nm) and OD (630 nm). TABLE5.1 The effect of Fengshiping on the TNF α nd IL-2 (X ± S)${{IL}\text{-}2\quad{activity}} = \frac{{{Sample}\overset{\_}{OD}} - {{Control}\quad\left( {{Culture}\quad{Fluid}} \right)\overset{\_}{OD}}}{\begin{matrix}{{{Standard}\quad{Sample}\quad\overset{\_}{OD}} - {{Control}\quad\left( {{Culture}\quad{Fluid}} \right)\overset{\_}{OD} \times}} \\{{activity}\quad{of}\quad{the}\quad{standard}\quad{sample}\quad\text{(IU/ml)}}\end{matrix}}$ dose Mouse TNF IL-2 group (g/kg) number (pg/ml) (IU/ml)Control — 10 87.80 ± 14.63  26.30 ± 4.22  12 10 62.14 ± 13.13** 16.00 ±2.89** Fengshiping 24 10 58.60 ± 9.63**  18.80 ± 2.86** 36 10 54.40 ±10.88** 18.20 ± 2.86** Tripterygium 8 10 58.25 ± 10.32** 16.00 ± 2.88**hypoglaucum (Levl.) Hutch. cyclophosphane 0.02 10 42.20 ± 9.57**  10.10± 3.00***P < 0.05,**P < 0.01

According to the data in Table 5.1, it suggested that the Fengshipinghave a obvious inhibiting effect on the TNF α. On the dosage of 12 g/kg,the medicine had showed a obvious inhibiting effect. Along with theincrease of the dosage, the inhibiting effect increased. But thedosage-effect curve went gently. The Fengshiping had an obviousinhibiting effect on the IL-2, ut no dosage-effect relationship wasobserved.

5.2 The Effect of Fengshiping on the IL-1, IL-6

70 NIH mice, 18-22 g weight, half male and half female, were dividedinto 7 groups at random. All the groups were drenched with thecorrespondent medicines (fengshiping and the other medicines). Themedicines were drenched once a day for 10 days. 24 hours after the lastadministration, all the mice were killed and sampled the macrophage andspleen cells from the abdominal carvity. The IL-1 and IL-6 in thesamples were measured.

The Measurement of IL-1:

The macrophages in the abdominal carvity were sampled in the asepsiscondition. Then the samples were washed by the Hank's liquor for 2 timesand nonserum RPMI1640 liquor for 1 time. Then the clear samples wereadjusted to the 4×10⁶/ml cell suspension with 5% FCS-RPMI liquor. 1 mlof the suspension was added to the test tube and cultured at 37° C. for1 hour. The unadherent cells were abandoned. Then the cultured liquorwas added with 5% FCS-RPMI 1640 and LPS (10 ng/ml) to culture. The cellscultured in 5% CO₂ at 37° C. for 72 hours. During the course, thecultured cells were freezed and thawed for several times. The finalproduct was saved at 4° C. The C57 mice were sampled the thymus in theasepsis condition. Then the samples were prepared to the 1×10⁶/ml cellsuspension with 5% FCS-RPMI1640.

100 μl supernate separated from the frost thawing liquor and 100 μl cellsuspension of thymus were added into the 96-hole flat bottomcell-culture batten. Each sample was cultured in 3 holes and comparedwith the different dilutions standard rHIL-1 and the control sample(culture fluid). Each hole was added with 2 ng ConA and then the battenwas cultured in the 5% CO₂ at 37° C. for 72 hours. 14 hours before theend of the culture, each hole was added with 3H-TdR 0.1μ Ci. Thecultured cells were collected with multihead cell-harvesting apparatusand measured the cpm value.${{IL}\text{-}1{activity}} = {\frac{{{Sample}\overset{\_}{cpm}} - {{Control}\quad\left( {{Culture}\quad{Fluid}} \right)\overset{\_}{cpm}}}{{{Standard}\quad{Sample}\overset{\_}{\quad{cpm}}} - {{Control}\quad\left( {{Culture}\quad{Fluid}} \right)\overset{\_}{cpm}}} \times {activity}\quad{of}\quad{the}\quad{standard}\quad\left( {{ng}\text{/}{ml}} \right)}$

The Measurement of the IL-6:

The spleen cells were sampled in the asepsis condition. Then the sampleswere washed by the Hank's liquor for 2 times and nonserum RPMI1640liquor for 1 time. Then the clear samples were adjusted to the 2×10⁶/mlcell suspension with 5% FCS-RPMI liquor. 1 ml of the suspension wasadded to the round-bottom centrifuge tube. After adding the ConA (10ng/ml), the samples were cultured in the 5% CO₂ at 37° C. for 72 hour.

The MH60 cells, which grew depending on the IL-6 and were on thelogarithmic growth stage, were adjusted to the 1×10⁵/ml cell suspensionwith the 5% FC-RPMI1640.

The 96-hole flat bottom cell culturing batten was added with the MH60cell suspension on the quantity of 100 μl/hole and the culturingsupernate 25 μl/hole. Then the fluid in each hole was adjusted to the200 μl with the 5% FCS-RPMI 1640. Each sample was cultured with 3 copiesand compared with the different solutions standard rHIL-6 and the pureculturing fluid. The batten was cultured in 5% CO₂ at 37° C. for 72hours. 6 hours before the end of the culture, the samples werecentrifuged. Each hole was sucked out the supernate 110 μl and added theMTT 10 μl. The samples were kept at 37° C. for 3 hours. And then theywere measured the OD at the wavelength 570 nm and 630 nm. The final ODvalue=OD 570 nm-OD 630 nm. TABLE 5.2 The effect of Fengshiping on theIL-1, IL-6 (X ± S)${{IL}\text{-}6\quad{activity}} = \frac{{SampleOD} - {{Culturing}\quad{Fluid}\quad{ControlOD}}}{\begin{matrix}{{{Standard}\quad{SampleOD}} - {{Culturing}\quad{Fluid}\quad{ControlOD} \times}} \\{{Sample}\quad{Dilution}\quad \times {Activity}\quad{Of}\quad{The}\quad{Standard}\quad\text{(IU/ml)}}\end{matrix}}$ Dose Mouse IL-1 IL-6 Group (g/kg) number (ng/ml) (IU/ml)Control — 10 78.7 ± 7.1  94.6 ± 6.8  7.5 10 59.3 ± 4.9** 64.9 ± 4.8** 1510 53.3 ± 5.7** 60.5 ± 4.3** Fengshiping 30 10 54.4 ± 4.8** 56.0 ± 4.6**60 10  47.0 ± 16.6** 56.6 ± 6.1** Tripterygium 5 10 57.6 ± 4.7** 65.7 ±4.9** hypoglaucum (Levl.) Hutch. cyclophosphane 0.02  9 44.5 ± 7.7  49.6± 6.7**

Based on the data in the table 5.2, the Fengshiping had an abviousinhibiting effect on the macrophage in producing of IL-1 and spleen cellin producing IL-6. Along with the increase of the dosage, the effectenhanced too.

5.3 The Effect of Fenghsiping on the Plasma NO in the AA Rat

60 SD rats, 160˜220 g weight, half male and half female, were dividedinto 6 groups. The rats in the blank control group were injected the NS0.5 ml under the skin of the right postpede vola. Other rats wereinjected with the FCA 0.5 ml at the same place of the control group. 18days later, the AA model was built. Then the rats were drenched thecorrespondent medicines or the distilled water once a day for 5 days. 3groups were drenched the solution of Fengshiping on the high, middle andlow dilution. The positive group was drenched with GlucosidorumTripterygll Totorum. The blank control group and the model group weredrenched with the distilled water of the same volumn. 1 hour after thelast administration, each rat was sampled the blood from the abdominalaorta for 2 ml. The plasma of the blood samples were separated and savedat −70° C. for the measurement. The measurement of NO was done on thedirection of the NO reagent. 0.1 ml plasma was added in 0.6 ml reagent Cand 0.4 ml double distilled water. After the mixture shaken up, it wasadded in 0.1 ml reagent D and cultured on the ice for 60 min. Then itwas centrifuged at 12000 rpm for 2 min. The supernate was separated. 0.6ml supernate was mixed with 0.4 ml double distilled water and 0.1 mlreagent A, and then it was cultured in the ice-water for 15 min. Thenthe mixture was added in reagent B 0.1 ml and put at the roomtermperature for 1 hour. Then the new mixture was measured the OD at thewavelength 545 nm. Based on the OD value of the sample, the content ofNO was calculated on the standard curve. (See the result in table 5.3)TABLE 5.3 The effect of Fengshiping on the plasma NO level in the AA rat(X ± S) Dose Rat Content of NO y Group (g/kg) number (μmol/L) (y = Lgx)Control — 8 13.55 ± 1.11* 1.131 ± 0.032 AA model — 9 17.56 ± 4.15 1.235± 0.097 Fengshiping 12 7  9.83 ± 2.58**^(ΔΔ) 0.985 ± 0.087 Fengshiping24 7 10.12 ± 1.56**^(ΔΔ) 1.001 ± 0.067 Fengshiping 48 7 10.70 ±1.51**^(ΔΔ) 1.026 ± 0.062 Glucosidorum 0.006 7 15.25 ± 3.48 1.173 ±0.099 Tripterygll TotorumComparing to the model group *P < 0.05, **P < 0.01; comparing to theGlucosidorum Tripterygll Totorum ^(ΔΔ)P < 0.01

Based on the data in table 5.3, the NO level was higher in the modelgroup than in the blank control group. The Fengshiping had an obviouseffect on lowering the NO level in the AA rat. The GlucosidorumTripterygll Totorum had the similar effect but its effect was weakerthan that of the Fengshiping.

EXPERIMENTAL EXAMPLE 6 The Effect of Fengshiping on the T Lymphocyte,CD₄, CD₈ and NK Cells in the Mouse

6.1 The Effect of Fengshiping on the Transform of Lymphocyte in theNormal Mouse

80 NIH mice, half male and half female, were divided into 8 groupsrandomly and drenched with the correspondent medicines once a day for 10days. 24 hours after the last administration, all the mice were killedto sample the spleen cells aseptically. Then the samples were washed bythe Hank's liquor for 2 times and nonserum RPMI1640 liquor for 1 time.Then the clear samples were adjusted to the 2×10⁶/ml cell suspensionwith 5% FCS-RPMI liquor. The 96-hole flat bottom cell culturing battenwas added with the cell suspension on the quantity of 100 μl/hole. Eachsample was cultured with 3 copies. 2 holes were added in 2 ng ConA eachas the stimulating reagent. The other hole was not added in the ConA andkept as the control hole. The batten was cultured in 5% CO₂ at 37° C.for 72 hours. 14 hours before the end of the culture, each hole wasadded in 3H-TdR 0.1μ Ci. The cells were harvested by the multihead cellharvesting instrument and measured the cpm value. The average value wasadopted as the sample's cpm value. The average value and the stimulatingindex of the different groups were compared directly. The stimulatingindex was calculated as following:

See the result in table 6.1 TABLE 6.1 The effect of Fengshiping on thelymphacyto transformation induced by ConA in the mouse (X ± S)${{Stimulating}\quad{Index}} = \frac{{Stimulated}\quad\overset{\_}{cpm}}{{Control}\quad\overset{\_}{cpm}}$Dose Mouse Stimulating Group (g/kg) number cpm index Control — 10 20433± 3579  25.87 ± 3.06 7.5 10 13566 ± 1779** 27.29 ± 7.67 15 10 12708 ±1692** 18.04 ± 3.76 Fengshiping 30 10 12809 ± 2575** 16.17 ± 4.37 60 1012090 ± 1706** 19.05 ± 3.80 2.5 10 18038 ± 3359  17.11 ± 2.60Tripterygium 5 10 12081 ± 1039** 17.58 ± 4.37 hypoglaucum (Levl.) Hutch.Cyclophosphane 0.02 9  9922 ± 1145** 13.66 ± 2.28Comparing to the control group*P < 0.05,**P < 0.01

According to the data in table 6.1, it indicated that the Fengshipinghad an obvious inhibiting effect on the lymphocyte transformation andthere was a dosage-effect relationship.

6.2 The Effect of Fengshiping on the CD₄, CD₈ and NK Cells

The experiment was same to 5.1. 24 hours after the last administration,the spleen cell samples were made into the 2×10⁸/ml cell suspension with5% FCS-RPIM1640. The quantity of CD₄, CD₈, NK cells and the rate CD₄/CD₈were measured on the usural method.

The Measurement of CD₄ and CD₈

The spleen cell suspension 50 μl was added on the glass to made the cellsmear. The glass had been coated by the polylysine. The T cell of themouse was set as the positive control sample. The cell smear wasenveloped by the serum of the normal mouse after it was fixed by theacetone. Then the enveloped sample was added with the antibody of CD₄and CD₈ which were marked by the hominine biotin. It was incubated at37° C. for 2 hours. Then the sample was added with the avidin labeled byenzyme and put still for 10 min. After added with the substrate for 10min, the mixed sample was washed and dyed with the hematoxylin for 2min. Then the sample was dyhidrated with the grade-alcohol and envelopedwith gelatin-glycetrol. 200 cells in the smear were chosen as theresearch target under the high power microscope.${{Content}\quad{Of}\quad{Cell}} = {\frac{{Dyed}\quad{cell}\quad{number}}{200} \times 100\%}$The Measurement of the NK Cell:

The preparation of the EC cell: The spleen cells were sampled in theasepsis condition. Then the samples were washed by the Hank's liquor for2 times and nonserum RPMI1640 liquor for 1 time. Then the clear sampleswere adjusted to the 2×10⁸/ml cell suspension with 5% FCS-RPMI liquor.This cell suspension was used as the EC.

The preparation of the TC cell: The Yack-1 cells, which were sensitiveto the mouse NK cell and on the logarithmic growth phase, were adjustedto the 4×10⁴/ml cell suspension. It was the TC.

Measurement: EC and TC, 100 μl each were added in the 96-hole flatbottom cell culturing batten. Each sample was cultured with 3 copies andset 2 control samples: EC and TC. (EC control: EC100 μl+5% FCS RPMI 1640100 μl; TC control: TC100 μl+5% FCS RPMI 1640 100 μl). The samples werecultured in 5% CO₂ at 37° C. for 24 hours. 6 hours before the end of theculturing, the samples were centrifuged and sucked out 110 μl supernateeach hole. And then the holes were added in the MTT 10 μl. After put at37° C. for 3 hours, the mixed samples were measured the OD value at thewavelength of 570 nm and 630 nm. The OD of each hole=OD570 nm-OD630 nm.TABLE 6.2 The effect of Fengshiping on the CD4, CD8, NK cell (X ± S)${{Activity}\quad{Of}\quad{NK}} = {\left( {1 - \frac{{{Sample}\quad\overset{\_}{OD}} - {{EC}\quad{Control}\quad\overset{\_}{OD}}}{{TC}\quad{Control}\quad\overset{\_}{OD}}} \right) \times 100\%}$Group Dose (g/kg) Mouse number CD4 (%) CD8 (%) CD4/CD8 NK Control — 1020.80 ± 2.94  14.80 ± 2.49 1.42 ± 0.18 40.13 ± 4.89    12 10 19.14 ±2.91  13.43 ± 2.51 1.43 ± 0.08 31.94 ± 4.52**^(ΔΔ) Fengshiping 24 1017.30 ± 2.51** 12.00 ± 2.40 1.46 ± 0.16 35.36 ± 3.40*^(ΔΔ)  36 10 16.30± 2.50**  11.23 ± 2.94** 1.49 ± 0.20 31.06 ± 3.53**^(ΔΔ) Tripterygium 810 16.25 ± 2.25** 11.50 ± 2.45 1.44 ± 0.18 32.20 ± 2.00**   hypoglaucum(Levl.) Hutch. Cyclophosphane 0.02 10 11.50 ± 2.50**   4.10 ± 1.20** 2.91 ± 0.53** 23.10 ± 3.66**  Comparing to the control group*P < 0.05,**P < 0.01;comparing to the cyclophosphane^(ΔΔ)P < 0.01

According to the table 6.2, it ie was a relation between the dzosage andthe effect, but the dosage-effect curve was smooth. The effective dosageof Fengshiping on the inhibiting of CD₄ was 24 g/kg. The minimumeffective dosage on inhibiting the CD₈ was 36 g/kg. As the rate ofCD₄/CD₈, the Fengshiping had no obvious effect. Cyclophosphane had anobvious effect on the inhibiting of the both kind of cells, and theinhibiting effect on the CD₈ was very powerful, which could increase therate of CD₄/CD₈ magnificently.

As for NK cell, the Fengshiping had a remarkable inhibiting effect, butthe dosage-effect relationship was not certain. As the same while, thecyclophosphane had shown an obvious inhibiting effect on the NK cell. Onthe dosage of 20 mg/kg, the inhibitiong effect of cyclophosphane wassignificantly different from that of the Fengshiping on the 3 dosages:12, 24 and 36 g/kg.

6.3 The Effect on the Transformation and Function of the T Lymphacyto inthe AA Mouse

NIH mice, 20±2 g weight, were injected with 0.05 ml FCA under the skinof the right postpede vola to build the AA model. The mice in thecontrol group were injected 0.05 ml NS at the same place. 3 weeks later,after the AA model built, all the mice were drenched the correspondentmedicines once a day for 5 days. 5 days later, all the mice were sampledthe blood to make the blood smear. The smears were dyed by the esterase.Then the smears were observed under the oil immersion lens to calculatethe percent of the positive-dyed cells (it represented the content ofthe T cells in the blood). The mice were sampled the spleen cells in thecondition of anaesthesia and then the cell samples were prepared to thesingle cell suspension. The cell suspension was washed by PBS and thenits supernate were abandoned. The rest part was added with bloodcytolysate 4 ml. The mixed sample was shaked for 2˜3 min to solute theRBC. After the RBCs were destroyed, the sample was centrifuged toseparate and abandon the supernate. The sample without supernate waswashed by the luminescence lotion for 2 times. Then it was centrifugedto separate and abandon the supernate. In the next step, the sample wasadjusted to the 1×10⁶/ml cell suspension. Each tube was added with 50 μldiluted antibody of CD₄ and CD₈. Then the tubs were cultured at 4° C.for 1 hour. After the culture, the samples were washed with theluminescence lotion for 2 times and added in the fixing fluid 2 ml.After fixing, the samples were filtrated through the 400-mesh screen tothe FCA tube. The filtrated samples were analyzed by the flow cytometer(FCM). The result was shown in the table 6.3. TABLE 6.3 The effect ofFengshiping on the T cell in the AA mouse (X ± S) Dose ANAE+ CD4+ CD8Group (g/kg) (%) (%) (%) CD4+/CD8+ Control — 50.60 ± 4.25 26.13 ± 1.1615.56 ± 0.68 1.68 ± 0.03 AAmodel — 49.00 ± 4.22^(▴) 32.56 ± 2.87** 13.59± 1.03** 2.49 ± 0.16** Fengshiping 7.5 49.13 ± 4.03^(▴) 27.30 ±1.76##^(▴) 15.98 ± 1.11##^(▴) 1.71 ± 0.04##^(▴) 15 49.31 ± 3.29^(▴)27.96 ± 1.67##^(▴) 16.23 ± 1.27##^(▴) 1.73 ± 0.05##^(▴) 30 48.56 ±3.23^(▴) 26.75 ± 1.94##^(▴) 15.58 ± 1.29##^(▴) 1.72 ± 0.04##^(▴)Glucosidorum 0.012 48.88 ± 2.89^(▴) 27.88 ± 1.99##^(▴) 16.33 ±1.31##^(▴) 1.70 ± 0.03##^(▴) Tripterygll Totorumn = 8,comparing with the control group *P < 0.05, **P < 0.01;comparing with the model group #P < 0.05, ##P < 0.01;comparing with the control group ^(▴)P > 0.05

According to the data in table 6.3, there was no significant differencein the different groups on the ANAE positive cell. But in AA mouse, theincrease of the CD₄ was significant, while the decrease of CD₈ wassignificant too. So the rate of CD₄/CD₈ had a remarkable increase. Theresult indicated that the Fengshiping could adjust the CD₄, CD₈ andCD₄/CD₈ to the normal range.

EXPERIMENTAL EXAMPLE 7 The Effect of Fengshiping on the PhagocyticFunction of the Macrophage in the Mouse Abdominal Carvity

50 NIH mice, 18˜22 g weight, half male and half female, were dividedinto 5 groups and drenched with the correspondent medicine solutions onthe same volumn. The administration was once a day for 7 days. 1 hourafter the last administration, all the mice were injected with 0.2 ml10% chick RBC into the abdominal carvity. 4 hours later, all the micewere killed and sampled the fluid in the abdomincal carvity. The liquorsamples were dropped on the glass and counted the number of themacrophage which had phagocytized the CRBC and the number of the CRBC inone macrophage. (See the result in table 7) TABLE 7 The effect ofFengshiping on the CRBC phagocytosis function of the macrophage in ICRmouse abdominal cavity (X ± S) dose Mouse Percent phagocytosis group(g/kg) number of phagocytosis (%) index Control — 10 25.75 ± 9.40  1.28± 0.20 Fengshiping 27 10 33.20 ± 12.77 1.46 ± 0.36 Fengshiping 40.5 1035.20 ± 10.16 1.21 ± 0.20 Fengshiping 60.9 10 37.78 ± 20.14 1.53 ± 0.32dexamethasone 0.005 10  8.33 ± 10.13* 1.10 ± 0.18*P < 0.05

According to the table 7, the Fengshiping had no obvious effect on thephagocytosis function of the macrophage in the mouse abdominal cavity.

EXPERIMENTAL EXAMPLE 8 The Effect of Fengshiping on the Hyperfunction ofthe Capillary Permeability in the Mouse Abdominal Cavity

90 NIH mice, 18˜22 g weight, half male and half female, were dividedinto 9 groups and drenched with the correspondent medicine solutions ofthe same volumn. The medicines were drenched once a day for 3 days orjust 1 time. 1 hour after the last administration, each mouse wereinjected with 0.7% HAC-NS solution into the abdominal cavity. At thesame time, each mouse was injected with the 0.5% Evans blue-NS solutioninto the vessel on the dose of 0.1 ml/10 g. 30 min later; all the micewere killed by cervical disjoint. The abdominal cavity was opened andwashed by the 5 ml NS. The NS used was collected and adjusted to 8 ml bythe pure NS as the sample. The samples were centrifuged at 3000 rpm toget the supernate. The supernate was measured the OD at the wavelengthat 590 nm. (See the result in table 8) TABLE 8 The effect of Fengshipingon the hyperfunction of the capillary permeability induced by the aceticacid in the mouse abdominal cavity (X ± S) Dose Leakage of the tinctureGroup (g/kg) Administration Mouse number (OD) P value Control — — 100.29 ± 0.13 Fengshiping 27 qd × 1 10 0.26 ± 0.14 >0.05 Fengshiping 40 qd× 1 10 0.25 ± 0.10 >0.05 Fengshiping 60 qd × 1 10 0.25 ± 0.09 >0.05Control — — 10 0.28 ± 0.15 Fengshiping 27 qd × 3 10 0.25 ± 0.12 >0.05Fengshiping 40 qd × 3 10 0.18 ± 0.10 <0.05 Fengshiping 60 qd × 3 10 0.15± 0.13 <0.05 dexamethasone 0.15 qd × 3 10 0.11 ± 0.07 <0.01

According to the data in table 8, it indicated that Fengshiping couldobviously inhibit the hyperfunction of the capillary permeabilityinduced by the acetic acid in the mouse abdominal cavity if it wasdrenched for 3 days continuously. If the medicine was drenched for just1 time, the inhibiting effect was not obvious.

EXPERIMENTAL EXAMPLE 9 The Effect of Fengshiping on the PleuritisExudation and the Inflammatory Cell Aggregation Induced by theCarrageenan

The mice were divided into 5 groups at random and injected with 0.5%Evans blue NS solution into the caudal vein on the dosage of 0.1 ml/10g. Then the mice were injected with the 0.03 ml 1% carrageenan in theright chest cavity with the special syring niddle. 4 hours and 32 hoursafter the injection, the correspondent mice were killed and opend theobdominal cavity to expose the diaphragm. 2 ml of the lotion wereinjected to the chest cavity by 2 times with a 1 ml injector. The lotionwas collected and saved in a test tube. 20 μl of the lotion collectedwas added into the 400 μl WBC dilution. The WBC in the mixed dilutionwas counted under the microscope. The rest of the lotion was centrifugedat 3000 rpm for 10 min. The supernate of the lotion was measured the ODat the wavelength of 600 nm. The OD value of the sample should becorrected with the correspondent OD value of the pure lotion. (See theresult in table 9) TABLE 9 The effect of Fengshiping on the inflammatorycell aggregation induced by the carrageenan (X ± S) Dose WBC number(2 ×105) Tincture exudation (OD) Group (g/kg) 4 h 32 h 4 h 32 h Control —46.0 ± 6.9 16.0 ± 9.6 0.156 ± 0.066 0.109 ± 0.019 Fengshiping 27 26.8 ±4.5* 14.2 ± 8.0 0.121 ± 0.062 0.116 ± 0.031 Fengshiping 40.5 10.9 ±4.0** 17.3 ± 4.6 0.100 ± 0.048 0.153 ± 0.032 Fengshiping 60  8.0 ± 5.5** 6.6 ± 4.7* 0.129 ± 0.066 0.092 ± 0.051 dexamethasone 0.05 12.7 ± 10.2** 4.4 ± 4.0* 0.085 ± 0.045 0.063 ± 0.017*P < 0.05,**P < 0.01

According to the table 9, it indicated that the Fengshiping had anobvious inhibiting effect on the inflammatory cell aggregation. Theeffect was powerful at the early stage. The regression equation on thedata of the fourth hour was as following: y=44.13−2.01x, r=−0.9625. Theeffect on the late stage was weak. At the high dosage of 20 g/kg, themedicine could affect the aggregation of the WBC. But it had no obviouseffect on the pleuritis exudation.

EXPERIMENTAL EXAMPLE 10 Effect on Aggregation of Leucocyte in Rats' CMCSac

Sixty four SD rats, 150-180 g weight, half male and half female, wererandomly divided into 8 groups, which were drenched with the same volumeand different dosage of drug liquid once a day, lasting 3 days. A daybefore experiment, rats were injected with 20 ml 1% CMC solution intothe sac at the rat's back caused by 20 ml air injection before theexperiment. 3.5 hour and 7.5 hour later, 0.1 ml liquid in the sac wasextracted each time, and was colored in 0.01% brilliant cresyl bluesolution. leucocyte was counted in the sac liquor under microscope. Theresults showed in the table 10. TABLE 10 effect on leucocyte counts ofcarboxymethyl cellulose sac of rats with Fengshiping (X ± S) dosage ratsWBC count(×107/L) groups (g/kg) number 3.5 hrs 7.5 hrs control — 8  9.7± 4.2 57.7 ± 17.3 Fengshiping 27 × 1 8  8.5 ± 3.5 39.4 ± 16.5Fengshiping 40 × 1 8  8.7 ± 7.3 35.3 ± 23.2 Fengshiping 60 × 1 8  6.6 ±3.3 18.1 ± 8.6** Control — 8 10.97 ± 6.7 35.6 ± 11.2 Fengshiping 27 × 38  15.4 ± 9.7 38.6 ± 15.5 Fengshiping 40 × 3 8  4.8 ± 3.4** 18.4 ±12.2** Fengshiping 60 × 3 8  3.0 ± 2.8** 11.0 ± 9.2* cortisone 0.1 × 3 8  14.2 ± 8.0 41.7 ± 16.0 Control — 8  10.9 ± 3.0 41.3 ± 6.9 Fengshiping18 × 7 8  6.2 ± 3.0* 11.4 ± 6.4* Fengshiping 27 × 7 8  3.7 ± 1.7**  6.4± 3.1** Fengshiping 40 × 7 8  2.5 ± 1.9**  5.9 ± 3.9** cortisone 2 mg ×1 8  1.5 ± 0.7**  3.0 ± 1.0**Compared with control group **P < 0.01

According to the table 10, the Fengshiping could inhibit significantlyaggregation of leucocyte in the rats' CMC sac, and the inhibition showedapparent dosage-effect relation, which was stronger as administrationtime lasted. With administration of continuing seven days, wandering ofleucocyte could be inhibited significantly at dosage of 118 g/kg, at thesame time, there was also very strong inhibition with cortisoneinjection into the sac.

EXPERIMENTAL EXAMPLE 11 The Effect on Croton Oil-Induced Swelling in theEars of Mice

60 NIH mice with weight of 18˜22 g, male and female accounting for halfand half, were divided into 6 groups, which were drenched with the samevolume and different dosage of drug liquid or tragacanth liquid, once aday, lasting 3 days. 1 hour after the final administration, 2% crotonoil mixure of 0.02 ml was embrocated uniformly on the both sides of leftears of mice, and after 4 hours, the mice were snapped off its cervicalvertebra and put to death. The left and right ears were cut down, theninflammatory and control ears were weighted by certain means. Differenceof weight between left and right ears was the swelling extent of ears,results showing in table 11. TABLE 11 effect on croton oil-inducedswelling of the ears of mice with Fengshiping (X ± S) dosage rats Degreeof ears' inhibition Groups (g/kg) number swelling (mg) rate (%) P valueControl group — 10 44.38 ± 9.40  Fengshiping 27 10 39.05 ± 12.3312.00 >0.05 Fengshiping 40 10 36.65 ± 5.83  17.64 <0.05 Fengshiping 6010 34.91 ± 9.71  21.34 <0.05 dexamethasone 0.003 10 14.13 ± 5.75  68.16<0.01

It was seen from table 11, that Fengshiping had remarkable inhibition tocroton oil-induced swelling of the ears of mice, and had quantity-effectrelation, but which curve was gentle and smooth. There was significantinhibition effect at 13.5 g/kg of dosage.

EXPERIMENTAL EXAMPLE 12 Effect on Acetic Acid-Induced Twisting Reactionof Mice

60 Kuming mice with weight of 18˜22 g, male and female accounting forhalf and half, were randomly divided into 6 groups, which were drencheddifferent dosages of drug liquid or Xihuangqi solution. 1 hours afteradministration, 0.7% HAC saline of 0.2 ml was injected, sc, and the micewere placed in aquarium and oberved the latent period before thetwisting reaction of each mouse and the twisting times in 20 minutes,results showing in table 12: TABLE 12 The effect of Fengshiping onacetic acid-induced twisting reaction of mice (X ± S) dosage Rats Latenttime groups (g/kg) numbers Twisting times (minute) Control — 10 34.6 ±14.1 3.13 ± 0.80 Fengshiping 27 10 28.2 ± 5.76 3.82 ± 0.85 Fengshiping40 10 31.0 ± 18.4 3.86 ± 2.00 Fengshiping 60 10  20.7 ± 12.3* 3.95 ±1.42 Tripterygium 20 10 25.1 ± 11.9 3.60 ± 0.93 hypoglaucum (Levl.)Hutch. morphine 10 mg/kg 10 0.0 ± 0.0 0.00 ± 0.00 hydrochloride

It was seen from table 12 that large dose of Fengshiping could delay thelatent time before the RAC-induced twisting reaction and significantlyreduce the twisting times in 20 minutes, which indicated Fengshiping hadthe effect of abirritation in some degree.

EXPERIMENTAL EXAMPLE 13 Effect on Hemorheology of AA Rats

Each of SD rats, 180±20 g weight, were injected intracutaneously with0.05 ml Freund's complete adjuvant on the right back foot metatarsal,and developed into adjuvant arthritis models. Each rats of negtivecontrol group were injected intracutaneously with 0.05 ml salin on theright back foot metatarsal. Three weeks after models built, the ratswere divided into model group, large, middle, small dosage group,negtive control group and positive control group which was administeredwith Glucosidorum Tripterygll Totorum. The rats were drenched once aday, lasting 5 days, 1 hours after administration for the last time, and3 ml blood was taken from abdominal aorta of rats and placed into testtube with 1% heparin as decoagulant, in which the whole blood viscositywas measured at shear rate of 230, 115, 46, 23, 11.5, 5.75 S⁻¹ withNXE-1 cone and plate viscometer. The plasma viscosity was measured withWTP-BII adjustable constant pressure capillary viscosimeter. Thehaematocrit, erythrocyte aggregation index was measured withcentrifugation method of packed cell volume. The rigidity index wascalculated from the above-mentioned data. All the results showed intable 13. TABLE 13 Effect on hemorheology of adjuvant arthritis modelrats (X ± S) Fengshiping Fengshiping Fengshiping GlucosidorumTripterygll Groups Control group Model group (30 g/kg) (15 g/kg) (7.5g/kg) Totorum (6 mg/kg) whole blood viscosity (mPa · s) 230S-1  4.43 ±0.09  4.92 ± 0.15**  4.56 ± 0.09##  4.49 ± 0.11##  4.54 ± 0.16##  4.66 ±0.28# 115S-1  5.17 ± 0.25  5.81 ± 0.19**  5.33 ± 0.09##  5.32 ± 0.10## 5.16 ± 0.14##  5.60 ± 0.48# 46S-1  6.84 ± 0.11  7.20 ± 0.18**  6.56 ±0.13##  6.59 ± 0.09##  6.67 ± 0.14##  6.70 ± 0.48# 23S-1  8.10 ± 0.15 8.23 ± 0.38  7.95 ± 0.22  7.93 ± 0.12  7.97 ± 0.14  8.02 ± 0.14 11.5S-1 9.35 ± 0.08  9.78 ± 0.10**  9.40 ± 0.08##  9.45 ± 0.10##  9.30 ± 0.133 9.31 ± 0.12## 6.5S-1 11.03 ± 0.14 12.66 ± 0.31** 11.21 ± 0.21## 11.29 ±0.19## 11.60 ± 0.40## 11.42 ± 0.52## Plasma viscosity 1.158 ± 0.0321.248 ± 0.040** 1.161 ± 0.011## 1.154 ± 0.023## 1.156 ± 0.018## 1.158 ±0.029## (mPa · s) corpuscular 46.13 ± 2.31 41.33 ± 1.12** 45.10 ± 2.39##44.33 ± 1.52## 45.71 ± 1.04## 46.03 ± 3.59## volume (%) erythrocyte 2.49 ± 0.032  2.58 ± 0.083*  2.46 ± 0.066#  2.49 ± 0.094#  2.44 ±0.048##  2.45 ± 0.091# aggregation index rigidity index 6.155 ± 0.5367.127 ± 0.557** 6.506 ± 0.558 6.525 ± 0.146 6.394 ± 0.200# 6.621 ± 0.883Compared with negative control group *P < 0.05, **P < 0.01;compared with model control group #P < 0.05, ##P < 0.01

According to the table 13, hemorheology of AA rats were changedsignificantly. The whole blood and plasma viscosity increased,haematocrit decreased, aggregation index and rigity index of erythrocyteincreased. The Fengshiping could make the above-mentioned indexes ofhemorheology improved significantly.

Pharmacological effects of Fengshiping have been proved by theabove-mentioned experiments. Many important pharmacological effects ofFengshiping had favorable dosage-effect relation, which implied the besttherapeutic effectiveness might be obtained by adjusting the drug dosageat clinical work.

The clinical studies on Fengshiping were carried on in China, Japan andAustrilia. Theses studies were operated according to internationalcriterion related disease classification about diagnosis, therapy andcurative effect. By using the Fengshiping capsules Sololy, its effectiverate was around 94%, and its remarkable effective rate was around 60%.It could improve the symptoms such as morning stiffness, swelling andpain and so on and the related items. The results showed in table 14˜21.TABLE 14 Compared effect of treatment group with control group remissionNo- Notable Effective (clinical table Effec- No effect rate Groups Casesrecovery) effect tive effect rate (%) (%) Treat- 32 5 14 11 2 59.3893.74 ment group Control 30 3 10 12 5 43.33 83.33 group

TABLE 15 Influence of IgG, IgA and IgM (X ± S) IgG IgA IgM Groups casespre- post- pre- post- pre- post- Normal 32 12.45 ± 1.48 2.37 ± 1.00 1.58± 0.59 Treatment 32 16.92 ± 3.49 14.17 ± 1.39** 3.65 ± 1.03 2.39 ±1.18** 1.89 ± 0.88 1.48 ± 1.01 group Control 30 17.03 ± 4.12 15.14 ±2.21** 3.45 ± 1.86 2.32 ± 1.75** 2.03 ± 0.95 1.76 ± 1.28Comparing with pre-treatment **P < 0.01

TABLE 16 Influence of C3 and C4(X ± S) C3 C4 groups cases pre- post-pre- post- normal group 32 0.62 ± 0.13 0.14 ± 0.15 Treatment group 321.88 ± 0.72 1.25 ± 0.66** 0.48 ± 0.12 0.26 ± 0.06*  Control group 302.13 ± 0.64 1.56 ± 0.62** 0.40 ± 0.16 0.25 ± 0.07**Comparing with before therapy *P < 0.05, **P < 0.01

TABLE 17 Influence of ESR and CRP (X ± S) ESR CRP Groups cases pre-post- pre- post- Normal 32 8.37 ± 5.26 4.12 ± 1.88 Treatment 32 66.58 ±9.01 30.31 ± 6.53**  13.35 ± 6.67 8.86 ± 3.34*  control 30 73.33 ± 9.0935.83 ± 11.61** 14.21 ± 6.29 9.04 ± 3.15**Comparing with pre-treatment *P < 0.05, **P < 0.01

TABLE 18 Compared with power of gripping pre- and post-treatment (X ± S)Treatment Group Control Group groups pre- post- pre- post- Grippingpower of left 39.13 ± 20.24(15) 80.47 ± 34.61**(15) 24.00 ± 17.63(21)55.15 ± 23.27**(21) hands (mmHg) Right hands 35.85 ± 22.46(15) 85.32 ±36.32**(15) 22.80 ± 12.32(21) 58.17 ± 20.59**(21)Comparing with pre-treatment *P < 0.05, **P < 0.01

TABLE 19 Influence of arthrosis swelling and pain and morning stiffnesstime (X ± S) Treatment Group Control Group Items pre- post- pre- post-arthrosis swelling and pain  5.79 ± 0.52  3.14 ± 0.83*  5.56 ± 2.15 3.92 ± 0.26* morning stiffness time 50.33 ± 6.47 20.24 ± 3.27** 48.75 ±8.34 27.50 ± 3.78** (minute)Comparing with pre-treatment *P < 0.05, **P < 0.01

TABLE 20 Influence of RF changing to negative RF negative Pre- Post-Rate of negative Groups Cases treatment treatment turnaround (%)Treatment 32 24 11 54.2 group Control 30 18 10 44.4 group

Not only had significant effects, but also Fengshiping can make itemssuch as SIL-2R, STNF, SIL-6R in plasma decrease, results showing in theTable 21. TABLE 21 inflence of main indes such as SIL-2R, STNF α ndSIL-6R (X ± S) SIL-2R(u/ml) STNF R1(ng/ml) SIL-6R(ng/ml) groups Casespre- post- pre- post- pre- post- Normal 32 299 ± 68 1.56 ± 0.48 72.05 ±18.26 (n = 32) (n = 24) (n = 22) Fengshiping 15 683 ± 189 381 ± 157**2.87 ± 0.66 1.75 ± 0.54** 136.18 ± 28.57 90.15 ± 20.12** Control 10 765± 203 412 ± 167** 2.63 ± 0.72 2.38 ± 0.39  148.21 ± 30.31 99.02 ±26.70** (n = 8)Comparing with pre-treatment **P < 0.01

It was proved that the above-mentioned results on invention could berealized on the ways as following.

PRACTICE EXAMPLE 1

-   -   Epimedium brevicornum Maxim. 2222 g    -   Tripterygium hypoglaucum (Levl.) Hutch. 2222 g    -   Lycium barbarum L. 1111 g    -   Cuscuta chinensis Lam. 1111 g

Four herbs hereinbefore, Tripterygium hypoglaucum (Levl.) Hutch. was cutinto pieces, extracted for three times after 13, 10, 10-fold added in,each time lasting 1 hour; Epimedium brevicornum Maxim was cut intosegments, extracted three times after 15, 10, 10-fold water was addedin, each extraction lasting 1 hour; Lycium barbarum L. was crushed toraw material, and immersed in 20-fold water of 80° C. for 1 hour;Cuscuta chinensis Lam. was crushed to raw powder, immersed in 31-foldwater of 80° C. for 1 hour; decoction fluid or immersion fluid of fourherbs were filtrated repectively, poured across macropore polymericadsorbent column, eluted with 70% ethanol. When the color of effluentbecame deep significantly, eluent was commenced to collect; when thecolor of effluent became very weak, elution collection was ended. Eluentof each herb was recycled to get ethanol. Then the fluid without alcoholwas concentrated, dried to get the finally extractive drug powder;officinal starch was blended with the four kinds of drug powder to 200g, mixed up uniformly and encapsuled into 1000 capsules. Each capsulewhich was prepared with the invented method thereof, was composed of 0.2g drugs extractive and contained at least 2.0 mg of icariine C₃₃H₄₀O₁₅.The regular dosage is: oral administration, three times every day, threecapsules every time.

PRACTICE EXAMPLE 2

-   -   Tripterygium hypoglaucum (Levl.) Hutch. 2000 g    -   Epimedium brevicornum Maxim. 2000 g

Two herbs hereinbefore, Tripterygium hypoglaucum (Levl.) Hutch. were cutinto pieces, extracted three times after 13, 10, 10-fold added in, eachtime lasting 1 hour; Epimedium brevicornum Maxim. was cut into segments,extracted three times after 15, 10, 10-fold water was added in, eachextraction lasting 1 hour; decoction fluid of herbs were filtratedrepectively, poured across macropore polymeric adsorbent column, elutedwith 70% ethanol, when the color of effluent became deep significantly,eluent was commenced to collect; when the color of effluent became veryweak, elution was over. Eluent of each herbs was recycled to getethanol, concentrated, dried, finally extractive drug powder wasobtained; officinal starch was blended with the extractive drug powder,and mixed up uniformly, loaded to 1000 capsules. Each capsule which wasprepared with the inventive method thereof, is composed of 0.2 g drugsextractive, contains at least 2.0 mg of icariine C33H40O15. regulardosage is: oral administration, three times every day, three capsulesevery time.

PRACTICE EXAMPLE 3

-   -   Tripterygium hypoglaucum (Levl.) Hutch. 2000 g    -   Epimedium brevicornum Maxim. 2000 g    -   Lycium barbarum L. 1000 g

Tripterygium hypoglaucum (Levl.) Hutch. were cut into pieces, extractedthree times after 13, 10, 10-fold added in, each time lasting 1 hour;Epimedium brevicornum Maxim. was cut into segments, extracted threetimes after 15, 10, 10-fold water was added in, each extraction lasting1 hour; Lycium barbarum L. was crushed to raw material, and immersed in20-fold water of 80° C. for 1 hour; decoction fluid or immersion fluidof four herbs were filtrated repectively, poured across macroporepolymeric adsorbent column, eluted with 70% ethanol, when the color ofeffluent became deep significantly, eluent was commenced to collect;when the color of effluent became very weak, elution was over. Eluent ofeach herbs was recycled to get ethanol, concentrated, dried, finallyextractive drug powder was obtained; officinal starch was blended withthe extractive drug powder, and mixed up uniformly, loaded to 1000capsules. Each capsule which was prepared with the inventive methodthereof, is composed of 0.2 g drugs extractive, contains at least 2.0 mgof icariine C₃₃H₄₀O₁₅. Regular dosage is: oral administration, threetimes every day, three capsules every time.

PRACTICE EXAMPLE 4

-   -   Tripterygium hypoglaucum (Levl.) Hutch. 2000 g    -   Epimedium brevicornum Maxim. 2000 g    -   Cuscuta chinensis Lam. 1000 g

Tripterygium hypoglaucum (Levl.) Hutch. were cut into pieces, extractedthree times after 13, 10, 10-fold added in, each time lasting 1 hour;Epimedium brevicornum Maxim. was cut into segments, extracted threetimes after 15, 10, 10-fold water was added in, each extraction lasting1 hour; Cuscuta chinensis Lam. was crushed to raw powder, immersed in31-fold water of 80° C. for 1 hour; decoction fluid or immersion fluidof the herbs were filtrated repectively, poured across macroporepolymeric adsorbent column, eluted with 70% ethanol, when the color ofeffluent became deep significantly, eluent was commenced to collect;when the color of effluent became very weak, elution was over. Eluent ofeach herbs was recycled to get ethanol, concentrated, dried, finallyextractive drug powder was obtained; officinal starch was blended withextractive drug powder, and mixed up uniformly, loaded to 1000 capsules.Each capsule which was prepared with the inventive method thereof, iscomposed of 0.2 g drugs extractive, contains at least 2.0 mg of icariineC₃₃H40O15. Regular dosage is: oral administration, three times everyday, three capsules every time.

PRACTICE EXAMPLE 5

-   -   Tripterygium hypoglaucum (Levl.) Hutch. 2000 g    -   Cuscuta chinensis Lam. 1000 g

Tripterygium hypoglaucum (Levl.) Hutch. Were cut into pieces, extractedthree times after 13, 10, 10-fold added in, each time lasting 1 hour;Cuscuta chinensis Lam. was crushed to raw powder, immersed in 31-foldwater of 80° C. for 1 hour; decoction fluid or immersion fluid of theherbs were filtrated repectively, poured across macropore polymericadsorbent column, eluted with 70% ethanol, when the color of effluentbecame deep significantly, eluent was commenced to collect; when thecolor of effluent became very weak, elution was over. Eluent of eachherbs was recycled to get ethanol, concentrated, dried, finallyextractive drug powder was obtained; officinal starch was blended withextractive drug powder, and mixed up uniformly, loaded to 1000 capsules.Dose of capsules administered every day, which was prepared with theinventive method thereof, was equivalent to dose of 30 g/day of crudedrugs.

PRACTICE EXAMPLE 6

-   -   Tripterygium hypoglaucum (Levl.) Hutch. 2000 g    -   Lycium barbarum L. 1000 g

Tripterygium hypoglaucum (Levl.) Hutch. were cut into pieces, extractedthree times after 13, 10, 10-fold added in, each time lasting 1 hour;Lycium barbarum L. was crushed to raw material, and immersed in 20-foldwater of 80° C. for 1 hour; decoction fluid or immersion fluid of theherbs were filtrated repectively, poured across macropore polymericadsorbent column, eluted with 70% ethanol, when the color of effluentbecame deep significantly, eluent was commenced to collect; when thecolor of effluent became very weak, elution was over. Eluent of eachherbs was recycled to get ethanol, concentrated, dried, finallyextractive drug powder was obtained; officinal starch was blended withextractive drug powder, and mixed up uniformly, loaded to 1000 capsules.Dose of capsules administered every day, which was prepared with theinventive method thereof, was equivalent to dose of 30 g/day of crudedrugs.

1. A pharmaceutical composition for treating rheumatism, characterizedin that, it is made from the following materials: Tripterygiumhypoglaucum (Levl.) Hutch. Epimedium brevicornum Maxim. Lycium barbarumL. Cuscuta chinensis Lam., Cuscuta australis R. Br. Wherein thematerials must be composed of Tripterygium hypoglaucum (Levl.) Hutch andone or two or three other herbs in the rest 3 herbs.
 2. Thepharmaceutical composition according to claim 1 made from the followingmaterials: Tripterygium hypoglaucum (Levl.) Hutch. 1-4 part by weightEpimedium brevicornum Maxim. 1-4 part by weight Lycium barbarum L. 1-4part by weight Cuscuta chinensis Lam., Cuscuta australis R. Br. 1-4 partby weight.
 3. The pharmaceutical composition according to claim 1 madefrom the following materials: Tripterygium hypoglaucum (Levl.) Hutch. 2part by weight Epimedium brevicornum Maxim. 2 part by weight Lyciumbarbarum L. 1 part by weight Cuscuta chinensis Lam., Cuscuta australisR. Br. 1 part by weight.
 4. The pharmaceutical composition according toclaim 1, characterized in that, it can be made from the correspondeffective constituents of the materials above-mentioned as followingthat Epimedium brevicornum Maxim. can be replaced by any one or morethan one among icariine, deuteron-icariine I, deuteron-icariine II andglyc-icariine A; Tripterygium hypoglaucum (Levl.) Hutch can be replacedby diterpenoids, triterpenoids and alkaloids compound thereof, andLycium barbarum L. and Cuscuta chinensis Lam. can be replaed by flavonecontained thereof.
 5. A method of preparing the pharmaceuticalcomposition according to claim 1, 2 or 3, characterized in that, itincludes the processes under-mentioned: The raw herbs are weighed, andEpimedium brevicornum Maxim. and Tripterygium hypoglaucum (Levl.) Hutch.were cut into pieces respectively; including raw material or crushedpowder of Lycium barbarum L. and Cuscuta chinensis Lam., four herbshereinbefore, were extracted with 0-95% ethanol at 10-98° C.respectively or combinatively for continuing 1-4 times. Ethanol wasrecycled respectively or combinatively in extracted fluid, thenextraction was concentrated, dried, crushed, mixed uniformly orproportionally, manufactured to dosage form adopted in clinical work;Raw herbs were weighed: Epimedium brevicornum Maxim. and Tripterygiumhypoglaucum (Levl.) Hutch. were cut into pieces, boiled out in water forthree times respectively, and Lycium barbarum L. or Cuscuta chinensisLam. were immersed in water of 80° C.˜95° C. for 1-3 times respectively.Decoction or immersion fluids of three times of each herb were blendedrespectively, then mixture fluid was respectively poured throughcorresponding macropore polymeric adsorbent column. After absorption,resin column was washed with water until effluent became clear, then waseluted with 30-99.5% ethanol until color of effluent became deep. Theneluent was collected until color of eluent became from deep to very weakwhile ethanol liquid was forced out from the column with water. Eluentwas mixed with the ethanol liquid. The weight of total eluent was 1-8fold of the herbs; eluent of each herbs was recycled, concentrated tospecific gavity of 1.10 respectively, then extractive of every herbswere obtained by respective or combinative spray drying, which weremixed uniformly and proportionally, manufactured to dosage form adoptedin clinical work.
 6. A method of preparing the pharmaceuticalcomposition according to claim 1, 2 or 3, characterized in that, it canbe made into any dose forms adopted in the clinical work such as hardgelatin capsule, soft capsule, tablet, granule and injection.
 7. Amethod of preparing the pharmaceutical composition according to claim 1,2 or 3, characterized in that, it includes the processesunder-mentioned: Tripterygium hypoglaucum (Levl.) Hutch. were cut intopieces, extracted three times after 13, 10, 10-fold added inrespectively, each time lasting 1 hour; Epimedium brevicornum Maxim. wascut into segments, extracted three times after 15, 10, 10-fold water wasadded in respectively, each extraction lasting 1 hour; Lycium barbarumL. was crushed to raw material, and immersed in 20-fold water of 80°C.-95° C. for 1 hour; Cuscuta chinensis Lam. was crushed to raw powder,immersed in 31-fold water of 90° C. for 1 hour; decoction fluid orimmersion fluid of four herbs were filtrated repectively, poured throughWLD or D101 or other type of macropore polymeric adsorbent column,eluted with 70% ethanol, when the color of effluent became deepsignificantly, eluent was commenced to collect; when the color ofeffluent became very weak, elution was over. Eluent of each herbs wasrecycled to get ethanol, concentrated, dried, finally extractive drugpowder was obtained; which were mixed uniformly and proportionally,manufactured to dosage form adopted in clinical work.
 8. The use of thepharmaceutical composition according to claim 1, 2 or 3 in themanufacture of a medicament for treating the rheumatoid and rheumatoidarthritis.
 9. The use of the pharmaceutical composition according toclaim 1, 2 or 3 in the manufacture of a medicament for treating thesystemic lupus erythematosus.
 10. The use of the pharmaceuticalcomposition according to claim 1, 2 or 3 in the manufacture of amedicament for treating the chronic nephritis, crohn's disease and leprareaction and the other autoimmune disease.